Carbohydrates are known to exert an important role in several biological and physiological pathways. It has been found, in fact, that they may contribute to the protein stability or resistance to degradation, to the regulation of intracellular signalling and targeting of glycoproteins to membranes or cell organelles, and to have implications in immunological recognition [see, as a general reference, Varki, A.; Glycobiology 1993, 3, 97-130, Bertozzi, C. L., Kiessling, L. L.; Science 2001, 291, 2357-64, Angeloni, S.; Glycobiology 2005, 15 (1), 31-41)].
In addition to that, several studies and research projects have discovered that carbohydrates and derivatives thereof, for instance glycans, take also part in some neoplastic modifications of cells. In fact, [as reported by Hakomori, S.; Chapter 4 in Montreuil, J., Vliegenthart, J. F. G., Schachter, H. (editors) Glycoproteins and disease. Elsevier, Amsterdam, 1996 or by Kobata A.; Cancer cells and metastasis. Chapter 3 in Montreuil, J., Vliegenthart, J. F. G., Schachter, H. (editors) Glycoproteins and disease. Elsevier, Amsterdam, 1996 or by Kumamoto et al in Biochemical and Biophisical Research Communications 1998, 247, 514-7 or by Hakomori, S.; PNAS 2002, 99 (16), 10231-33], it has been observed that glycans, known to be present on cell membranes, usually exhibit structural modifications, for instance a high degree of branching in cancer cell membranes over normal cell membranes.
Likewise, besides any structural modification, it has also been observed an altered composition in membrane carbohydrates with respect to normal cells.
As an additional example of a saccharidic moiety playing a role in neoplastic processes there is, for instance, sialic acid (see The Merck-Index; XIII Ed.; No. 8558), a 9-carbon atom amino sugar being a member of neuraminic acid related compounds. There has been observed, in fact, that the high metastatic activity of certain tumoral forms may be well correlated with an increased concentration of sialic acid in cell membranes, thus yielding to a reduced adhesive capability of the extra-cellular matrix cells in metastatic phenomena (see, for a reference, Varki, A. et al Essentials of glycobiology. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. 1999).
Additionally, for instance as reported by Sames, D. et al in Nature 1997, 389, 587-91 or by Dwek, M. V. et al. (Clinica Chimica Acta 1998, 271, 191-202) or by Burchell et al (Glycobiology 1999, 9 (12), 1307-11) or by Taylor-Papadimitriou et al. (Biochim. Biophys. Acta 1999, 1455, 301-13) or by Hanisch et al (Glycobiology 2000, 10 (5), 439-49) or by Schuman, J. et al (Glycoconjugate Journal 2000, 17, 835-48) within complex saccharidic compounds, a highly glycosilated glycoprotein like altered MUC1, which shows a high number of O-glycosidic residues and that provides a protective layer on epithelial surfaces and it is involved in cell-cell interactions, signalling, and metastasis (see, for a reference Parry S, Silverman H S et al., Biochem. Biophys. Res. Commun. 2001; 11; 283(3): 715-20), seems to contribute to the metastatic potential of the neoplastic cells.
Though numerous molecular alterations have been found to be associated with each single developing or progressing phase of a tumor, the exact relationship among them is not yet completely understood.
The phenotypic alterations in saccharides found to characterize a neoplasia, are generally known as Tumor Associated Carbohydrate Antigens (hereinafter shortly referred to as TACAs).
The epitopes Gal-β-1→3-GalNAc-α-O.Ser/Thr and GalNAc-α-O.Ser/Thr (see the chemical structures below), commonly referred to as TF and Tn respectively, are among the most widespread known TACAs.

As from the above reported formulae, the Tn immunodeterminant group is a GalNAc residue α linked to the hydroxyl group of serine or threonine, which appears in the amino-terminal region of glycoproteins. In its turn, Gal-β-1→3-GalNAc-α-O.Ser/Thr (TF antigen) is then formed by the action of the enzyme β-galactosyltransferase, which adds a galactose residue, taken from a Gal-nucleotide, to a Tn residue.
The occurrence of these antigens in normal tissues has been investigated in order to verify their specificity toward pathological tissues (see, for example, Cao, Histochem. Cell Biol. 1996, 106, 197-207).
Moreover, TF and Tn antigens were found to be over-expressed in immunoreactive form, to a significant extent, in about 90% of carcinoma tissues (see, for a reference, Springer, G. F.; Science 1984, 224, 1198-206) and their relative proportion in human carcinomas frequently correlates with the carcinoma's aggressiveness itself.
The results of several studies also suggested that at the early stages of the metastatic process, TF and Tn antigens both play a key role in adhesion phenomena between the tumoral cells and the adjacent normal cells (see, for a reference, Kischikawa et al, in Jpn. J. Cancer Res. 1999, 90, 326-32).
The isolation and structural identification of carbohydrate antigens specifically over-expressed in cancer cells, and thus of TF and Tn, represents a first step for the development of a strategy for the therapy and diagnosis of neoplastic diseases: the carbohydrate-based immunotherapy and immunodiagnosis (see, for instance, Allen in J. Aim Chem. Soc., 2001, 123, 1890-7).
It is well known in the art that immunotherapy involves the stimulation of the natural body defenses, i.e. immunosystem, against harmful agents such as microorganisms, pollutants, chemicals, foods and the like. Therefore, it has been envisaged that cancer cells, characterized by an altered expression of TACAs on their membrane, might represent the target of the human or animal immunosystem.
There was found that the stimulation of the immunosystem through suitable tumoral antigens such as, for example Tn or TF antigens, could be a promising therapeutic tool against tumor.
Toyokuni et al. (see, for a reference, Bioorg. Med. Chem. 1994, 2, 119-32) describes the preparation of totally synthetic carbohydrate vaccines by the use of monomeric, dimeric and trimeric Tn antigens further conjugated with Ovine Serum-Albumin, Starbust® dendrimer and tripalmitoyl-S-glycerylcysteinylserine (P3CS).
Lo-Man et al. (see, for example, J. Immunol. 2001, 166, 2849-54) discloses the development of a fully synthetic immunogen called multiple antigenic glycopeptide and based on a dendrimeric lysine core with four arms.
Kuduk et al. (see, for a reference, J. Am. Chem. Soc. 1998, 120, 12474-12485) describes the use of KLH, BSA and a lipopeptide (pam) for the clusterization of Tn and TF antigens, while Dziadek et al. (Angew. Cem. Int. Ed. 2005, 44, 7624-7630) discloses a similar approach for the preparation of compounds comprising TACAs and BSA and MUC1 peptides.
Kagan et al. (see, for a reference, Cancer Immunol. Immunother. 2005, 54, 424-430) describes the tests made on several Tn conjugates: Tn monosaccharide, Tn(c) prepared on a triple threonine backbone and Tn prepared on a partially or fully glycosylated MUC1 backbone.
Among the major limitations of the above reported conjugated compounds, however, is the fact that when used as vaccines, they lead to the production of poor quantities of the desired antibodies and, also, that an aspecific mixture of the same is finally recovered (for example, see Grigalevicius et al, Bioconjugate Chemistry, 2005, 16, 1149-1159).
We have now found novel conjugated compounds of TF and Tn antigens that may be used as vaccines in the therapy against tumors. The conjugated compounds of the invention, in addition, allow to overcome the above drawbacks of the TACAs conjugated compounds of the prior art.